SUMO (Small Ubiquitin-like Modifier) is a ubiquitin-like protein that can covalently modify a large number of cellular proteins. SUMOylation is a dynamic process that is mediated by activating (E1), conjugating (E2), and ligating (E3) enzymes and readily reversed by a family of SUMO-specific proteases (SENP). We showed that inactivation of the SENP1 gene causes embryonal lethality in mid- gestation in mice as a result of severe fetal anemia stemming from deficient erythropoietin (Epo) production. SENP1 controls Epo production by regulating the stability of hypoxia-inducible factor 11 (HIF11) in hypoxic condition. During normoxia, HIF11 is hydroxylated in two critical proline residues by a family of oxygen sensing enzymes. Proline hydroxylation is important for HIF11 to binds to its ubiquitin ligase complex, von Hippel-Lindau protein VHL/elongin B/C complex, leading to ubiquitination and proteasomal degradation. We showed that hypoxia induced rapid SUMOylation of HIF11, which allowed it to bind to VHL in a hydroxyl-proline-independent manner, also leading to ubiquitination and proteasomal degradation. SENP1 reverses SUMOylation of HIF11, reduces binding to VHL, and consequently stabilizes HIF11. Thus, SENP1 plays a critical role in regulating HIF11 stability during hypoxia. Since HIF11 regulates multiple, critical downstream genes, such as Epo and VEGF, it has the potential to regulate erythropoiesis and angiogenesis. In preliminary results, we showed that accumulation of SUMOylated HIF11 peaked at four hours after exposure to hypoxia and later declined. This corresponded to an early phase of SUMOylation, most likely by an increase in SUMO E3 activity and a later phase of de-SUMOylation, most likely mediated by an increase in SENP1 activity. In this proposal, we will examine the role of SUMOylation and de-SUMOylation in HIF11 regulation by hypoxia and its downstream effects, focusing on angiogenesis. AIM 1: To identify the E3 responsible for hypoxia-induced SUMOylation of HIF11. AIM 2: To study how SENP1 is regulated by hypoxia. AIM3: To determine whether SENP1 regulates angiogenesis during development and in the tumor environment. These studies should provide novel insights into how the de-SUMOylation pathway regulates the hypoxic response.